1991 Amer. Soc. Microbiol. Ann. Meetings, Dallas, TX

Transter RNA.Mutation in the Halophilic Archaea.

DANIEL T. NIEUWLANDT1, ELIZABETH S. HAAS2, TRACI BALTRUS1, DAVID ARMBRUSTER1, and CHARLES J. DANIELS1*

1Ohio State Univ., Columbus, OH; 2 Indiana Univ., Bloomington, IN.

We have investigated two enzymes of the tRNA maturation pathway in Haloferax volcanii, intron endonuclease and RNaseP. In vitro studies on the endonuclease have revealed that this enzyme recognizes a defined structural element present at the exon-intron boundaries. Absent is a requirement for mature tRNA structure, a defined characteristic of the eukaryotic endonuclease. To determine if similar recognition properties exist in vivo, we have developed an expression vector for H. volcanii. Using this vectorl we have introduced modified forms of the H. volcanii tRNA Trp intron containing gene and analyzed their in vivo maturation. As predicted, alterations of the structure at the exon-intron boundaries have profound effects on cleavage efficiency.

We have also characterized the RNA component of the H. volcanii RNaseP complex. This RNA is similar to its eubacterial counterparts both in structure and sequence. Sequence analysis of five related halobacterial RNaseP RNAs has revealed conserved and highly variable regions. Like eubacterial RNaseP RNAs, these RNAs can potentially form two pseudoknot structures. Whether these RNAs have catalytic activity in the absence of protein remains an active question.